WE GO WHERE NO OTHERS CAN

SINGLE MOLECULE miRNA COUNTING BY A PCR FREE METHOD

Hugh Ilyine

CEO Destina Genomics Ltd.

hugh@destinagenomics.com

20th February 2020 Manchester Biomarker Conference

DESTINA is a company focused ondelivering high value, high performanceRNA diagnostic assays for diagnosticsand drug development

Small (19 - 25 nt) non-coding RNA molecules that regulate

gene expression at the post-transcriptional level

Specific for pathologies

Altered before clinical symtons appear

Detection from biological fluids

Current methods - Blurred pictures

From analogue to digital

THE RNA BIOMARKER DETECTION DREAM

• No isolation, concentration of targets needed

• Direct detection in all biological fluids, without isolation, concentration of targets –RT-qPCR free

• No refrigeration of samples needed between prep and test, or during transport

• No errors, perfect reproducibility from lab to lab, without concern for inhibitors

• Absolute quantification, not relative quantification, no normalisers needed

• Run in the same machine as protein biomarkers, same sample, same time.

• Fast, cost effective, low technicity required

Taqman PCR and Next Generat ion Sequenc ing

Limitations for microRNA detection:

• Small molecules give errors when

using PCR

• High costs and time-to-results

• Lack of accuracy and lab to lab

reproducibility

PLATFORM AGNOSTIC

Multiple platforms possible• Bead based - colourimetric• Array based – colourimetric, flourescent• Mass based• Magnetic Particle based

High specificity, sensitivity depends on platform used

WHY DESTINA AND AN SMCX-PRO?

• Quantify RNA directly in specimens

• No extraction or amplification required

• Absolute quantification vs relative

• Stabilise RNA at point of collection

• Fast and reliable results

Key – how to dissociate labelled heteroduplex from beads

Hybridization

25ºC

From 10 mL

serum/plasma

DESTINA

Stabiltech

buffer

60 min

1. Bead separation*

2. Washings*

3. DCL Rxn

60 min

1. Bead separation*

2. Washings*

3. Alexa Labelling (20 min)

4. Dissociation (4 min)

30 min

Samples can be stored

and/or shipped at room

temperature (up to 14 days)

* Bead separation and washing steps

done in automatized equipment.

** 96 well-plate formatTarget RNA

Read

RESULTSRobust Cal ibrat ionCur ves

LoD = 1 .5 pM

Sample = 10 uL

N=3

DIL I pat ients serum

14.0 pM to 46 .9 pM

Tr ipl icate per pat ient

Liver-specific microRNA biomarker

Liver disease miRNA signature Expression

Alcoholic liver diseasemiR-122 (acute alcohol, microsteatosis) ↑

miR-122, miR-155 (chronic alcohol, macrosteatosis) ↑

Hepatitis CmiR-122, miR-34a, miR-155, miR-125b, miR-146a, miR-21 ↑

Hepatitis B miR-192, miR-122 ↑

Hepatocellular carcinoma

miR-21 miR-16, miR-199a, miR-122, miR-223, miR-885-5p ↑

Drug overdose miR-122, miR-192 ↑

Degradation of miR-122 is blockedby using DESTINA StabiltechTM

buffer

Stabilises samples at collectionpoints, during transport andsample holding and handlingwithout any refrigeration needed

Multiple miR-122 isomiRs in DILI are less presentin serum from healthy patients

The canonical form of miR-122 was not the mostabundant in both healthy and DILI patients

Sample degradation over time produces moreisomiRs

microRNA degradation was substantiallyincreased if DILI occurs

NGS

A) SYBR Green PCRB) Taqman PCRC) DCL assayD) Virtual patient analysis

DESTINA Genomics Ltd. DESTINA Genómica S.L7-11 Melville St, Edinburgh EH3 7PEUnited Kingdom

+44 (0) 162 082 2611

info@destinagenomics.com

Avenida de la Innovación 1, 18016 GranadaSpain

+374 660 159 779

spain@destinagenomics.com